The long-term goal of this research is to devise methods to improve the prevention and treatment of periodontitis and serious dental infections by gaining an increased understanding of the biology of the black-pigmented Bacteroides. This group of anaerobic gram-negative bacteria was chosen for study for two main reasons. First, they are important causes of periodontitis and serious oro-facial infections that complicate dental procedures. Second, these bacteria have recently become increasingly resistant to penicillin and tetracycline, the standard antibiotics used to treat dental infections. This project will employ the current techniques of molecular biology with the objectives of uncovering the mechanisms of antibiotic resistance and the virulence factors of the black-pigmented Bacteroides. The rationale for these studies is based on the recent development in this laboratory of a novel system of genetic analysis of the intestinal Bacteroides fragilis group of anaerobes. This recombinant DNA technology can now be applied to study the oral anaerobes. The specific aims of this project are: 1) to characterize the genetic mechanisms of tetracycline and penicillin resistance in the black-pigmented Bacteroides by defining and cloning the resistance genes, 2) to develop a system of genetic transfer, using shuttle plasmids, between E. coli and the black-pigmented Bacteroides, and 3) to develop techniques for transposon mutagenesis in these Bacteroides. The latter two aims are essential for the development of a system for genetic manipulation which will involve cloning and mutagenesis of important drug resistance and virulence genes. This molecular approach to the mechanisms of virulence and antibiotic resistance of the black-pigmented Bacteroides will provide a broader understanding of the biology of these bacteria, and is likely to yield information necessary to achieve breakthroughs in the prevention and treatment of dental infections.